Journal of Analytical Toxicology

WU, Alan H.B.; Feng, Yue-Jin; Pajor, Audrey; Gornet, Terrie G.; Wong, Shan S.; Forte, Elaine; Brown, Jill

doi: 10.1093/jat/21.3.181pmid: 9171199

A total of 2259 urine samples were assayed for lysergic acid diethylamide (LSD) using radioimmunoassay (RIA, Coat-a-Count, Diagnostics Products) and a premarket cloned enzyme donor immunoassay (CEDIA, Boehringer Mannheim). Urine samples were obtained from patients admitted to the emergency room, patients in drug rehabilitation programs, and adults and juveniles in criminal probation programs. An overall incidence of positive results was 0.80% for CEDIA (500-pg/mL cutoff) and 0.89% and 0.18% for RIA at cutoffs of 250 and 500 pg/mL, respectively. Of the CEDIA-positive samples, only 17 and 11% were positive by RIA at 250 and 500 pg/mL, respectively, whereas among RIA-positive samples, only 10% of those > 250 pg/mL and only 25% of those > 500 pg/mL were positive by CEDIA. Moreover, only 2 of 25 of samples positive by one of these screening assays were confirmed by gas chromatography-mass spectrometry (GC-MS). It is likely that discrepancies in results between immunoassays are due to differences in antibody specificities used to detect LSD metabolites. In addition, immunoassays may be more sensitive than GC-MS for detecting LSD use as current confirmation assays are targeted towards detection of the parent drug only. The interpretation of results for LSD analysis must be made with knowledge of the limitations for each assay.

Kudo, Keiko; Jitsufuchi, Narumi; Imamura, Tohru

doi: 10.1093/jat/21.3.185pmid: 9171200

A selective, sensitive, and reliable method was devised to determine concentrations of amitriptyline and its major metabolite, nortriptyline, in human plasma using high-performance liquid chromatography (HPLC) combined with UV and particle beam mass spectrometry (PBMS). Amitriptyline and nortriptyline were effectively extracted in a three-step solvent extraction procedure. Imipramine was used as the internal standard (IS). Amitriptyline, nortriptyline, and the IS were clearly separated by HPLC on a silica column with the mobile phase of acetonitrile-0.1M ammonium acetate (94:6, v/v). The calibration curves were linear in the concentration range of 10–1000 ng/g for both compounds with UV and PBMS detections. The lower limits of detection were 5 ng/g for amitriptyline and 10 ng/g for nortriptyline with UV detection and 2 ng/g for amitriptyline and 5 ng/g for nortriptyline with PBMS detection. The absolute recoveries were 58% for amitriptyline and 47% for nortriptyline at a concentration of 50 ng/g. This method proved most useful in accurately identifying amitriptyline and nortriptyline in tissues from an autopsied individual.

Brady, Thomas C.; Yang, T.J.; Hyde, Walter G.; Kind, Albert J.; Hill, Dennis W.

doi: 10.1093/jat/21.3.190pmid: 9171201

A two-step kinetic enzyme-linked immunosorbent assay was developed to detect the presence of flunixin in the urine of greyhound dogs. The assay system was developed using polyclonal antiflunixin antisera, a rabbit albumin-flunixin conjugate adsorbed onto polystyrene microtiter strips, and flunixin reference standards for calibration. The assay parameters were optimized and the performance characteristics were determined. The quantitative intra- and inter-run precisions (%CV) of the analysis of replicate (n = 10) flunixin-spiked urine samples were 9.9–12.5% and 10.2–13.6%, respectively. The linear dynamic range was 1–100 ng/mL, and the quantitative accuracy, as determined by calculation of percent error of measured flunixin in flunixin-spiked drug-free greyhound urine, was −16% to +14% over this range. The I50 of the ELISA was 17.3 ng/mL. The limit of detection was 25 ng/mL in greyhound urine. The reactivity in the assay system relative to flunixin (100%) was 147% for flunixin glucuronide, 25% for clonixin, and 5% for niflumic acid. The ELISA was capable of detecting total flunixin for up to 72 h in dogs administered flunixin at 0.55 mg/kg orally and up to 96 h in a dog that was administered flunixin at 1.0 mg/kg orally.

Sarkar, Pratibha; McIntosh, John M.; Leavitt, Randy; Gouthro, Heather

doi: 10.1093/jat/21.3.197pmid: 9171202

Nimesulide is a nonsteroidal anti-inflammatory drug recently detected in equine blood and urine samples taken at the race track. The detection of the drug in a blood sample led to the identification of an unknown thin-layer chromatographic (TLC) spot in track urine samples as a metabolite of nimesulide. Characterization of the unknown TLC spot and comparison with the synthesized compound shows that the unknown TLC spot is a previously unreported equine metabolite of nimesulide. The metabolite was identified as resulting from the reduction of the nitro group on nimesulide to an amino group. This reduced nitro metabolite (4-amino-2-phenoxy-methanesulfonanilide) is a major metabolite of nimesulide in the equine.

Queiroz, R.H.C.; Dreossi, S.A.C.; Carvalho, D.

doi: 10.1093/jat/21.3.203pmid: 9171203

An original, simple, specific, and rapid high-performance liquid chromatography assay for the determination of dapsone and monoacetyldapsone in human plasma is presented. The procedure consists of extracting the drugs and the internal standard (phenacetin) from 1 mL plasma with ethyl ether at alkaline pH. A liquid chromatograph equipped with a reversed-phase 5-µm C8 analytical column was used. The drugs were eluted with a mixture of water and methanol (70:30, v/v). Flow rate and wavelength were set at 1 mL/min and 286 nm, respectively. The precision, linearity, and limit of quantitation of the method were within acceptable limits. The method was considered adequate and has been used for the determination of the acetylator phenotype in population studies.

Mannaert, Erik; Tytgat, Jan; Daenens, Paul

doi: 10.1093/jat/21.3.208pmid: 9171204

A qualitative screening technique was developed for the detection of 2-amino-5-chloropyridine, a newly identified decomposition product of zopiclone and metabolites after alkaline hydrolysis of urine samples. The method was elaborated using a standard operation procedure (Merck Tox Screening System), combining a solid-phase extraction with reversed-phase high-performance liquid chromatography and diode array detection. The limit of detection, expressed as zopiclone concentrations in spiked urine, was found to be 0.5 µg/mL. Field urine samples from a volunteer were positive for 2-amino-5-chloropyridine until 16 h after ingestion of one therapeutic dose of Imovane (7.5 mg zopiclone).

Cagle, Joan C.; McCurdy, H. Horton; Pan, Y. Mary; Ayton, Kim J.; Wall, William H.; Solomons, Everett T.

doi: 10.1093/jat/21.3.213pmid: 9171205

The Microgenics CEDIA DAU (EIA) and the Abbott AxSym system (FPIA) cannabinoids assays were evaluated for their combined effectiveness in the analysis of cannabinoids in whole blood. Blood samples were treated with acetone, evaporated, and reconstituted, and the supernatant was analyzed by the EIA cannabinoids assay. Blood samples determined positive by EIA were then treated with acetonitrile and sodium sulfate, and the resultant protein-free supernatant was analyzed using the FPIA cannabinoids assay. A total of 98 blood samples determined to be presumptively positive by both EIA and FPIA were further analyzed for the presence of 11-nor-carboxy-Δ9-tetrahydrocannabinol-9-carboxylic acid (THCCOOH). All 98 blood samples could be confirmed for the presence of THCCOOH by gas chromatography-mass spectrometry (GC-MS) at concentrations greater than the 10 ng/mL cutoff. The GC-MS results were found to correlate significantly better with those of the FPIA cannabinoids assay (r = 0.75) than with EIA (r = 0.22). Procedures for the rapid analysis of whole blood for cannabinoids using CEDIA DAU reagents and the AxSym system are presented.

Jenkins, Amanda J.; Levine, Barry; Locke, J. Laron; Smialek, John E.

doi: 10.1093/jat/21.3.218pmid: 9171206

Alprazolam is one of the most widely prescribed benzodiazepines in the United States. It is generally considered a safe and effective drug for the treatment of anxiety disorders and panic attacks. Few overdoses that are due to the sole ingestion of alprazolam have been reported. This paper documents a fatality due to alprazolam intoxication and describes the distribution of alprazolam and an active metabolite, α-hydroxyalprazolam, in tissues obtained at autopsy. Qualitative identification of the drugs was achieved by full-scan gas chromatography-mass spectrometry, and quantitative analysis was performed by high-performance liquid chromatography. High concentrations of alprazolam were found in all specimens analyzed, but the metabolite was detected only in subclavian blood, urine, bile, and liver. A postmortem heart blood alprazolam concentration of 2.1 mg/L is the highest reported in the literature to date.

Cosbey, S.H.; Carson, D.J.L.

doi: 10.1093/jat/21.3.221pmid: 9171207

A fatal case attributed to amlodipine intoxication is presented. The deceased was a 15-year-old girl who allegedly ingested 14 10-mg Istin tablets. Amlodipine concentration in peripheral blood was determined (2.7 mg/L) and was compared with published therapeutic and toxic data for amlodipine and some other dihydropyridine calcium channel-blocking agents. Amlodipine concentrations in liver, blood, and stomach contents were also determined.

De Baere, S.; Meyer, E.; Dirinck, I.; Lambert, W.; Piette, M.; Van Peteghem, C.; De Leenheer, A.

doi: 10.1093/jat/21.3.223pmid: 9171208

A fatality that was due to the ingestion of the halogenated solvent trichloroethylene is presented. The decedent was a 43-year-old male who was found dead at his home. Screening of the blood and stomach contents with the enzyme multiplied immunoassay technique and radioimmunoassay demonstrated the presence of ethanol, amphetamine-like compounds, caffeine, cotinine, and acetaminophen. These compounds were present in toxicologically irrelevant concentrations as confirmed by thin-layer chromatography, high-performance liquid chromatography, and gas chromatography (GC). The Fujiwara reaction was performed on all available matrices, and it revealed the presence of chlorinated hydrocarbons in high concentrations. A specific GC method with electron capture detection allowed the quantitation of trichloroethylene and its metabolites trichloroethanol and trichloroacetic acid in different matrices. GC with Fourier-transform infrared detection was used for the confirmation of the identity of trichloroethylene.